Through affinity labeling of phosphorylase kinase with adenine nucleotide analogs we hope to gain information about which subunit(s) of the enzyme have catalytic capacity. By measuring the rate of inactivation toward different protein substrates caused by the affinity labels we will determine if the enzyme has more than one type of catalytic site, and if so, which substrates are converted at which sites. We will attempt to apply this type of analysis to the autophosphorylation reaction also. An additional goal will be to determine the properties of the autophosphorylated phosphorylase kinase. For instance, does autophosphorylation affect its Ca2 ion-dependency or its action on other protein substrates? Also, what is the rate and extent of phosphate incorporation into the individual subunits during autophosphorylation under a variety of conditions.